-
Research ArticleNew Collagen-Dextran-Zinc Oxide Composites
forWound Dressing
Georgeta Psunica-Panea,1,2 Anton Ficai,3 Minodora Maria Marin,4
S, tefania Marin,4
Msdslina Georgiana Albu,4 Vlad Denis Constantin,1,2 Cristina
Dinu-Pîrvu,5 Zina Vuluga,6
Mihai Cosmin Corobea,6 and Mihaela Violeta Ghica5
1Surgery Department, “Carol Davila” University of Medicine and
Pharmacy, Dionisie Lupu 37, 020022 Bucharest, Romania2Surgery
Department, “Sfântul Pantelimon” Emergency Clinical Hospital,
340-342 S, oseaua Pantelimon, 021659 Bucharest, Romania3Faculty of
Applied Chemistry andMaterial Science, Politehnica University of
Bucharest, 1-7 Polizu Street, 011061 Bucharest, Romania4Collagen
Department, Leather and Footwear Research Institute, 93 Ion
Minulescu Street, 031215 Bucharest, Romania5Department of Physical
and Colloidal Chemistry, Faculty of Pharmacy, “Carol Davila”
University of Medicine and Pharmacy 6,Traian Vuia Street, 020956
Bucharest, Romania6Polymer Department, National Institute for
Research & Development in Chemistry and Petrochemistry,202
Splaiul Independent,ei Boulevard, 060021 Bucharest, Romania
Correspondence should be addressed to Mihai Cosmin Corobea;
[email protected]
Received 29 September 2015; Accepted 28 December 2015
Academic Editor: Paulo Cesar Morais
Copyright © 2016 Georgeta Păunica-Panea et al.This is an open
access article distributed under theCreative
CommonsAttributionLicense, which permits unrestricted use,
distribution, and reproduction in anymedium, provided the
originalwork is properly cited.
The goal of this paper was the design, development, and
characterization of some new composites, based on collagen and
dextran asnatural polymers and zinc oxide as antimicrobial, to be
used in wound healing. Collagen hydrogels with various
concentrations ofdextran and zinc oxide were investigated in terms
of rheological analysis. The spongious composites, obtained by
freeze-drying ofhydrogels, were evaluated by morphology (SEM),
water uptake, and biological (enzymatic biodegradation) analysis.
All the resultswere strongly influenced by the nature and
concentration of composite components. Based on the performances of
the hydrogels,stationary rheometry, porous structure, morphology,
and biological behavior, the antimicrobial spongious composite
based oncollagen and dextran with 50% ZnO were the most promising
for future applications in wound dressing and a biomaterial
withhigh potential in skin regeneration.
1. Introduction
Wound healing is a significant problem for health-caresystems
worldwide, accounting over 1.5% of the world pop-ulation [1]. The
most affected by chronic wounds, as ulcers,are elderly and diabetic
people.Moreover, an untreated or notcorrectly treated wound can
lead to large area of necrosis andto systemic infection [2]. To
avoid such complications, thebest solution is the use of
antibacterial biomaterials to treator prevent infection of the
tissues.
Collagen is one of themost used polymers in biomaterialsfield,
due to its excellent properties in
biocompatibility,biodegradability, with well-established structure,
biologicprofile, and in vivo response [3]. It is a bioactive
medical
device used in different types of injuries (varicose
ulcer,burns, wounds, opened surgery, etc.) as haemostatic
andmedical dressing [4]. Being a natural protein, collagen
itselfcannot heal the infected tissue because bacteria use it as
asubstrate [5–7]. Another natural polymer, dextran, a
polysac-charide was proven to stimulate wound healing, control
theproliferation of bacteria, and affect the metabolism of
tumorcells, smooth muscle cells, and endothelial cells [8].
Dextran-based hydrogel containing chitosan microparticles
loadedwith growth factors [9] and silk fibroin
nanofibrousmaterialswith dextran [10] were also successfully used
in woundhealing, but some simpler solutions were not exploited
yet.
In order to induce antimicrobial activity, the
polymericscaffolds have to be more bioactive by decoration with
Hindawi Publishing CorporationJournal of NanomaterialsVolume
2016, Article ID 5805034, 7
pageshttp://dx.doi.org/10.1155/2016/5805034
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2 Journal of Nanomaterials
antimicrobials like antifungals, antivirals, antiparasitics,
ornonpharmaceutical (like essential oils). Zinc oxide
(ZnO)waswidely related to exhibit antimicrobial activity and
higherstability than organic molecules [11, 12]. Also, it was used
toaccelerate the healing of both chronic and acute wounds
[13]because of its epithelialization and bacteriostatic
properties.ZnO represents today one of the most reliable choices
inobtaining composites with potential applications in woundscare
[14, 15].
The aim of this study is to develop new simple solutionsfor
wound dressings based on collagen and dextran. Thescaffold was
designed by using dextran for wound healingand ZnO for
antimicrobial properties. The systems areprepared by lyophilization
method in order to obtain effi-cient absorbent properties for wound
dressings and porousstructures. The composites in form of hydrogels
were eval-uated by rheological analysis and the spongious forms
wereinvestigated by water uptake, biodegradability in
collagenasesolution, and SEM.
2. Materials and Methods
2.1. Materials and Reagents. Type I fibrillar collagen gelhaving
a concentration of 2.46% (w/w) was extracted fromcalf hide using
the technology currently available at theResearch-Development
Textile Leather National InstituteDivision Leather and Footwear
Research Institute—CollagenDepartment [3]. Dextran from Leuconostoc
spp. (Mw 15,000–25,000) was purchased from Fluka (USA) and zinc
oxidenanopowder (
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Journal of Nanomaterials 3
Table 2: The determination coefficients’ values for different
rheological models tested at 37∘C.
Rheological models/hydrogels G1 G2 G3 G4Casson 0.9832 0.9481
0.9418 0.9397Bingham 0.9233 0.8432 0.8308 0.8168Ostwald-de Waele
0.9838 0.9904 0.9913 0.9954Herschel-Bulkley 0.9959 0.9936 0.9931
0.9976
Table 3: Herschel-Bulkley model fitting parameters for collagen
hydrogels with different concentrations of ZnO.
Rheological parameters/hydrogels G1 G2 G3 G4Yield stress (Pa)
4.707 9.704 13.559 16.406Consistency index (Pa⋅s𝑛) 5.251 22.282
30.920 33.787Flow index 0.445 0.287 0.218 0.212
2.6. In Vitro Degradation by Collagenase. In order to
investi-gate the enzymatic degradation of collagen scaffolds,
massloss was monitored as function of exposure time to a
colla-genase solution. Pieces of 1 × 1 cm collagen composites
wereimmersed in a collagenase solution and incubated at 37∘C.At
predetermined intervals, the swollen pieces were removedfrom the
collagenase solution and weighed. The percent ofdegradation was
calculated using the following equation:
%weight loss =𝑊𝑖−𝑊𝑡
𝑊𝑡
× 100, (6)
where 𝑊𝑖is the initial weight and 𝑊
𝑡is the weight of the
samples after a time 𝑡. All the samples were studied
intriplicate.
2.7. Scanning Electron Microscopy (SEM). SEM analyses
wereperformed on a HITACHI S2600N electron microscope, onsamples
coveredwith silver layer for each collagen composite.
3. Results and Discussion
The influence of ZnO concentration on upward flow curvesplotted
as shear stress as a function of shear rate for hydrogelsG1 ÷ G4 is
presented in Figure 1.
The upward rheograms presented in Figure 1 indicatedthat the
hydrogels G1 ÷ G4 showed a non-Newtoniancharacter, the shear stress
increase with shear rate increase.
Table 2 summarizes the values obtained for determina-tion
coefficients by fitting the experimental data to variousrheological
models ((1)–(4)) described in Section 2.
As can be seen from Table 1, the Herschel-Bulkley modelbest
fitted the rheological data shear stress as a function ofshear
rate.
The dependence of ZnO concentration on the flowdescriptors
characteristic to Herschel-Bulkley model is givenin Table 2.
The data presented in Table 3 show that the hydrogelsG1 ÷ G4
exhibit a pseudoplastic behaviour with yield stressfacilitating
their flow and allowing their good manipulation[16, 22]. The values
of flow index between 0.212 and 0.445indicate a high degree of
pseudoplasticity, especially forG2–G4. The presence of ZnO in
formulation determined
G1G2
G3G4
248 12 16 2040Shear rate (s−1)
0
16
32
48
64
80
Shea
r stre
ss (P
a)
Figure 1: The up flow curves for collagen hydrogels with
differentZnO concentrations tested at 37∘C.
an increase of consistency index and yield stress, the
highestvalues being recorded for the hydrogel containing a
con-centration of 75% ZnO related to collagen dry substance.The
values of the aforesaid descriptors increase with ZnOconcentration,
more obvious for the hydrogels with 25% and50% concentration. Thus,
the presence of a lower concen-tration of ZnO (G2) determined the
doubling of yield stressvalue and also a marked increase of
consistency index value(about 4.24 times) compared to sample G1.
Moreover, thedoubling of ZnO concentration from 25% to 50%
determineda more important increase of the rheological parameters
forG3 related toG2 (40% for yield stress and 38% for
consistencyindex) compared to G4 related to G3, increase from 50%
to75%, respectively (21% for yield stress and 38% for
consistencyindex).
Similar patterns as recorded for G1 ÷ G4 were obtainedfor the
hydrogels G5 ÷ G8 with dextran, the flow curvesbeing also described
by the Herschel-Bulkley model. Theaddition of dextran in
formulations does not significantly
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4 Journal of Nanomaterials
0
9
18
27
36
45
M1 M2 M3 M4 M5 M6 M7 M8W
ater
upt
ake (
g/g)
Collagen composites
2 days4h
Figure 2: Water uptake for collagen composites.
modify rheological parameters specific to the above modelfor
collagen-zinc oxide hydrogels.
The results of the rheological analysis showed that
theconcentration of zinc oxide is the main factor influencingthe
flow properties of the designed hydrogels. Thus, besidethe own
pharmacological effect, zinc oxide concentrationsignificantly
affects the flow parameters.
The G1 ÷ G8 hydrogels were freeze-dried and spongiouscollagen
composites (M1 ÷ M8) were obtained and char-acterized by water
absorption, enzymatic degradation, andmorphology by SEM.
The water uptake capacity is a very important propertyfor an
ideal wound dressing in order to maintain a moistenvironment and to
keep the excessive exudates. Figure 2presents the water absorption
after 4 hours and 2 days for thecollagen composites obtained.
The control sample based on cross-linked collagen, M1,had higher
water uptake capacity, compared to the oneswhich contain dextran or
zinc oxide. The water uptakedecreased as zinc oxide concentration
increases from 25% to75% (to collagen content). This behavior could
be due to thepresence of an increased concentration of zinc oxide
whichfavors a more dense composite structure. The addition
ofdextran to the samples M1 ÷M4 decreases the water
uptakecapacity.
The results were confirmed also by SEM images (Figure 3)which
showed the compositional involvement on the spon-gious composites
morphology. According to SEM obser-vations all the scaffolds formed
a three-dimensional (3D)porous structure. The pores inside the
scaffolds were inter-connected and varied in a large range,
depending on zincoxide and dextran content. For M1 (reference
sample) theassociated morphology (Figure 3(a)) of the structure
sug-gests pore sizes between 80 and 270𝜇m, along with theincrease
of zinc oxide concentration the size of pores decreaseto 55–105 𝜇m
(Figure 3(b)). Moreover ZnO particles couldbe clearly seen on
collagen fibrils on SEM images. Dex-tran induced a more homogeneous
phase appearance, with
smaller pores, with sizes between 48 and 75𝜇m (Figure 3(c)).From
Figure 3(d) it can be seen that pores forms were moreuniform in the
presence of dextran and ZnO nanoparticlesadhered on collagen
fibrils (Figures 3(d) and 3(e)).
Both the zinc oxide and the dextran affected the finalporous
structures of the scaffolds. Materials obtained fromcollagen
hydrogel composites with dextran presented a more“compact”
structure and the pore diameter was much smallercompared to the
neat collagen ones.
In vitro biodegradation of collagen composites by collage-nase
solution was assessed to simulate the in vivo behavior ofcomposites
used as wound dressings. High degradation rateswere registered
(Figure 4) for collagen samples without zincoxide: over 50% after 4
hours and totally after 2 days.
The dextran content increased the resistance to collage-nase and
zinc oxide improved the overall stability of the sam-ples. The
relative collagen degradation content decreased forsampleswith 25%
and 50%ZnO.When large amounts of ZnO(75%)were used, the enzymatic
degradation slightly increasescompared with 50% ZnO sample. The
same tendency relatedto composition was observed also for the
collagen dextransystems.These results can be explained by the
higher amountof minerals which leads to a lower homogenous
compositephase; therefore the collagen network can be more
exposedto the enzymatic degradation (i.e., on the outer shell ofthe
inorganic particles). Among the composites studied, themost stable
ones at both 4 hours and 2 days were the onewith collagen, dextran,
and 50% ZnO. Moreover, taking intoaccount the flow analysis for the
corresponding hydrogels, theuse of ZnO concentration higher than
50% was not justifiedsince it does not lead to a marked increase of
the rheologicalparameters and to a higher stability for the porous
compositesdegradation. Considering the wound healing application,
theeffectiveness of the composite depends on the decreasingcollagen
rapid degradability profile for prolonged treatmentand healing
efficiency.
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Journal of Nanomaterials 5
(a) (b)
(c)
(d) (e)
Figure 3: SEM images of collagen composites: (a) M1 (×200); (b)
M4 (×200); (c) M5 (×200); (d) M7 (×500); and (e) M7 (×50000).
4. Conclusions
ZnO particles increased the pseudoplastic behavior of
thecomposites based on collagen in solution phase. Dextranpresence
did not significantly influence the rheological pro-files of the
hydrogels. ZnO particles increased the consistencyindexes and
reduced the flow indexes of solution phases.
The flow parameters indicated the Herschel-Bulkley modelfor
better describing the rheological behavior.
Water uptake ability for the hydrogel composites withdextran and
collagen was lower, in comparison with neatcollagen ones. ZnO
particles were able to reduce the wateruptake for both collagen and
collagen/dextran matrixes.
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6 Journal of Nanomaterials
M1 M2 M3 M4 M5 M6 M7 M8Collagen composites
2 days4h
0
20
40
60
80
100
120
Wei
ght l
oss (
%)
Figure 4: In vitro enzymatic degradation of collagen
composites.
SEM morphology indicated the decrease of pore sizesand an
increase in uniformity when ZnO was used. Theinorganic particles
were found in a dispersed state decoratingthe collagen and
collagen/dextran fibrils.
Using ZnO particles the in vitro degradation profile of
thecomposites can be adjusted by increasing the length of
thedegradation process (enzymatic assisted).The biodegradabil-ity
of the composites can be tailored by both dextran andZnOparticles.
But large amounts of ZnO particles (75%) can leadto disruption of
the composite phase distribution by exposingmore collagen to the
enzymatic process.
Based on the performances of the hydrogels stationaryrheometry
and of porous structures morphological and bio-logical
investigations, the antimicrobial spongious compositebased on
collagen and dextran with 50% ZnO could beselected for applications
to patientwounds, being a promisingbiomaterial in skin
regeneration.
Conflict of Interests
The authors declare no conflict of interests.
Acknowledgments
This work was supported by the “Parteneriate in
DomeniiPrioritare—PNII,” supported by MECS-UEFISCDI (Projectno.
155/2014) acronym GREENVET. This work receivedfinancial support
through the project entitled “CERO—Career Profile: Romanian
Researcher,” Grant no. POS-DRU/159/1.5/S/135760, cofinanced by the
European SocialFund for SectoralOperational
ProgrammeHumanResourcesDevelopment 2007–2013.
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